The association between human papillomavirus (HPV) and the development of cervical cancer has encouraged research in methods for HPV testing. The objective of this effort is to develop an in situ hybridization assay for detecting on Thin Prep slides the wide range of pathogenic HPV types infecting the cervix. ThinPrep technology is a liquid based filtration method that allows the cleansing and monolayering of cervical cells on slides. This cleansing process reduces blood, mucus, and nondiagnostic debris yielding highly discernible specimens for cervical screening. Thin Prep slides have improved the sensitivity in detection of abnormal cells, and reduced the number of cervical specimens with SBLB (Satisfactory But Limited By) status. In Phase I, we will develop an in situ HPV assay that utilizes ThinPrep slides, peptide nucleic acid probes, and the highly sensitive tyramide signal amplification system. In Phase II, this assay will be optimized with respect to methods of cell digestion, cell fixation, hybridization stringency conditions, chromogen detection, block and wash reagents. Thereafter, this assay will be used in a clinical setting to establish its potential in discriminating which women should be treated aggressively; i.e., requiring follow up colposcopy or biopsy. Phase III will focus on product development and clinical trials. PROPOSED COMMERCIAL APPLICATION: The HPV market is anticipated to reach $60,000,000 in the next five years.The commercialization of an in situ method capable of triaging patients with different cervical abnormalities will be an important first step in implementing a more cost effective and efficient approach for detecting and treating women at high risk for cervical cancer.